The present invention relates to novel isoxazolopyrimidinones, to processes for preparing them and to their use as medicaments, in particular as inhibitors of cGMP-metabolizing phosphodiesterases.
The publication Bull. Soc. Chim. Belg. (1994), 103 (5-6), 181-4 describes the synthesis of isoxazolo[4,5-d]pyrimidinones which are unsubstituted in the 5 position. Compounds possessing a phenyl radical or substituted phenyl radical in the 5 position are not described. There is no report of the described substances having a biological effect, in particular being able to inhibit phosphodiesterases.
The compounds according to the invention are potent inhibitors of cyclic guanosine 3xe2x80x2,5xe2x80x2-monophophate-metabolizing phosphodiesterases (cGMP-PDEs). In accordance with the nomenclature of Beavo and Reifsnyder (Trends in Pharmacol. Sci. 11, 150-155, 1990), these phosphodiesterases are the phosphodiesterase isoenzymes PDE-I, PDE-II and PDE-V.
An increase in the concentration of cGMP can lead to therapeutic, antiaggregatory, antithrombotic, antiproliferative, antivasospastic, vasodilatory, natriuretic and diuretic effects. It can exert an effect on the short-term or long-term modulation of vascular and cardiac inotropy, cardiac rhythm and stimulus conduction in the heart (J. C. Stoclet, T. Keravis, N. Komas and C. Kugnier, Exp. Opin. Invest. Drugs (1995), 4 (11), 1081-1100). Inhibition of the cGMP-PDEs can also strengthen erection. These compounds are therefore suitable for treating erectile dysfunction.
The present invention now relates to novel isoxazolidinones of the general formula (I) 
in which
R1 represents (C3-C8)-cycloalkyl or represents straight-chain or branched alkyl having up to 4 carbon atoms,
A represents radicals of the formulae 
in which
R2 and R7 are identical or different and denote hydrogen or straight-chain or branched alkyl having up to 4 carbon atoms,
R3, R4, R5 and R6 are identical or different and denote hydrogen, a 5- to 6-membered aromatic heterocycle having up to 3 heteroatoms from the series S, N and/or O or aryl having from 6 to 10 carbon atoms, where the ring systems are optionally substituted, up to three times, identically or differently, by hydroxyl, halogen, (C1-C5)-alkyl or (C1-C6)-alkoxy, or
R3, R4, R5 and R6 are identical or different and denote (C1-C6)-alkyl which is optionally substituted by aryl having from 6 to 10 carbon atoms and can, for its part, be substituted, up to 3 times, identically or differently, by (C1-C6)-alkyl, (C1-C6)-alkoxy, hydroxyl or halogen, or
R3 and R4 and/or R5 and R6, together with the nitrogen atom to which they are bonded, form radicals of the formulae 
where
R8 denotes hydrogen or (C1-C6)-alkyl, which is optionally substituted by hydroxyl, and
R9 denotes hydrogen or hydroxyl,
and the salts, N-oxides and isomeric forms thereof.
The compounds according to the invention can exist in stereoisomeric forms which either relate to each other as image and mirror image (enantiomers) or which do not relate to each other as image and mirror image (diastereomers). The invention relates to both the enantiomers or diastereomers or their respective mixtures. The racemic forms, as well as the diastereomers, can be separated into the stereoisomerically uniform constituents in a known manner.
The substances according to the invention can also be present as salts. Within the context of the invention, preference is given to physiologically harmless salts.
Physiologically harmless salts can be salts of the compounds according to the invention with inorganic or organic acids. Preference is given to salts with inorganic acids, such as, for example, hydrochloric acid, hydrobromic acid, phosphoric acid or sulfuric acid, or salts with organic carboxylic or sulfonic acids, such as, for example, acetic acid, maleic acid, fumaric acid, malic acid, citric acid, tartaric acid, lactic acid or benzoic acid, or methanesulfonic acid, ethanesulfonic acid, phenylsulfonic acid, toluenesulfonic acid or naphthalenedisulfonic acid.
Physiologically harmless salts can equally well be metal or ammonium salts of the compounds according to the invention. Particular preference is given, for example, to sodium, potassium, magnesium or calcium salts, and also to ammonium salts which are derived from ammonia, or to organic amines, such as, for example, ethylamine, di- or triethylamine, di- or triethanolamine, dicyclohexylamine, dimethylaminoethanol, arginine, lysine, ethylenediamine or 2-phenylethylamine.
(C3-C8)-Cycloalkyl represents cyclopropyl, cyclopentyl, cyclobutyl, cyclohexyl, cycloheptyl or cyclooctyl. Those which may be mentioned as being preferred are: cyclopropyl, cyclopentyl and cyclohexyl.
(C6-C10)-Aryl represents an aromatic radical having from 6 to 10 carbon atoms. Preferred aryl radicals are phenyl and naphthyl.
(C1-C6)-Alkyl represents a straight-chain or branched alkyl radical having from 1 to 6 carbon atoms. Those which may be mentioned by way of example are: methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl and n-hexyl. Preference is given to a straight-chain or branched alkyl radical having from 1 to 4 carbon atoms. Particular preference is given to a straight-chain or branched alkyl radical having from 1 to 3 carbon atoms.
(C1-C6)-Alkoxy represents a straight-chain or branched alkoxy radical having 1 to 6 carbon atoms. Those which may be mentioned by way of example are: methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, n-pentoxy and n-hexoxy. Preference is given to a straight-chain or branched alkoxy radical having from 1 to 4 carbon atoms. Particular preference is given to a straight-chain or branched alkoxy radical having from 1 to 3 carbon atoms.
Halogen generally represents fluorine, chlorine, bromine and iodine. Preference is given to fluorine, chlorine and bromine. Particular preference is given to fluorine and chlorine.
A 5- to 6-membered aromatic heterocycle having up to 3 heteroatoms from the series S, O and/or N represents, for example, pyridyl, pyrimidyl, pyridazinyl, thienyl, furyl, pyrrolyl, thiazolyl, oxazolyl or imidazolyl. Preference is given to pyridyl, pyrimidyl, pyridazinyl, furyl and thienyl.
Preference is given to compounds according to the invention of the general formula (I),
in which
R1 represents (C3-C6)-cycloalkyl or represents n-propyl,
A represents radicals of the formulae 
in which
R2and R7 are identical or different and denote hydrogen, methyl or ethyl,
R3, R4, R5 and R6 are identical or different and denote hydrogen, pyridyl, pyrimidyl, pyridazinyl, furyl, imidazolyl, thienyl, pyrryl or phenyl, where the ring systems are optionally substituted, up to 2 times, identically or differently, by hydroxyl, fluorine, chlorine, (C1-C4)-alkyl or (C1-C4)-alkoxy, or
R3, R4, R5 and R6 are identical or different and denote (C1-C4)-alkyl which is optionally substituted by phenyl which, for its part, can be substituted, up to 2 times, identically or differently, by (C1-C4)-alkoxy, fluorine or chlorine, or
R3 and R4 and/or R5 and R6, together with the nitrogen atom to which they are bonded, form radicals of the formulae 
where
R8 denotes hydrogen or (C1-C6)-alkyl which is optionally substituted by hydroxyl, and
R9 denotes hydroxyl,
and the salts, N-oxides and isomeric forms thereof.
Particular preference is given to compounds according to the invention of the general formula (I),
in which
R1 represents cyclopentyl or n-propyl,
A represents radicals of the formulae 
in which
R2 and R7 are identical or different and denote methyl or ethyl,
R3, R4, R5 and R6 are identical or different and denote hydrogen, pyridyl or phenyl, where the ring systems are optionally substituted, up to 2 times, identically or differently, by fluorine, methoxy or ethoxy, or
R3, R4, R5 and R6 are identical or different and denote (C1-C3)-alkyl which is optionally substituted by phenyl which, for its part, is substituted, up to 2 times, identically or differently, by methoxy or ethoxy, or
R3 and R4 and/or R5 and R6, together with the nitrogen atom, form radicals of the formulae 
where
R8 denotes hydrogen or (C1-C3)-alkyl or a radical of the formula xe2x80x94(CH2)2xe2x80x94OH, and
R9 denotes hydroxyl,
and the salts, N-oxides and isomeric forms thereof.
Very particular preference is given to the following compounds:
In addition, a process for preparing the compounds according to the invention of the general formula (I) was found, which process is characterized in that
compounds of the general formula (II) 
in which
R1 has the abovementioned meaning,
are initially converted, by reaction with compounds of the general formula (III)
Axe2x80x2xe2x80x94COxe2x80x94Clxe2x80x83xe2x80x83(III),
in which
Axe2x80x2 represents radicals of the formulae 
in which
R2 and R7 have the abovementioned meaning,
in inert solvents, where appropriate in the presence of a base, into the compounds of the general formula (IV) 
in which
Axe2x80x2 and R1 have the abovementioned meaning,
a cyclization is subsequently carried out, in the presence of POCl3 or of alkali metal hydroxide or alkaline earth metal hydroxide, to give the compounds of the general formula (V) 
in which
Axe2x80x2 and R1 have the abovementioned meaning,
and, in a further step, the compounds of the general formula (VI) 
in which
Axe2x80x2 and R1 have the abovementioned meaning,
are prepared by reaction with ClSO3H,
and, finally, are reacted with amines of the general formula (VII)
HNR10R11xe2x80x83xe2x80x83(VII),
in which
R10 and R11 encompass the abovementioned scope of meaning of R3, R4, R5 and R6.
The process according to the invention can be explained, by way of example, by means of the following formula schemes: 
Solvents which are suitable for the individual steps are the customary organic solvents which are not altered under the reaction conditions. These solvents preferably include ethers, such as diethyl ether, dioxane, tetrahydrofuran or glycol dimethyl ether, or hydrocarbons, such as benzene, toluene, xylene, hexane, cyclohexane or petroleum fractions, or halogenohydrocarbons, such as dichloromethane, trichloromethane, tetrachloromethane, dichloroethane, trichloroethylene or chlorobenzene, or ethyl acetate, dimethylformamide, hexamethylphosphoric triamide, acetonitrile, acetone, dimethoxyethane or pyridine. It is likewise possible to use mixtures of the solvents mentioned.
In general, the reaction temperatures can vary over a relatively wide range. In general, the temperatures employed are in a range of from xe2x88x9220xc2x0 C. to 200xc2x0 C., preferably of from 0xc2x0 C. to 70xc2x0 C.
In general, the process steps according to the invention are carried out under standard pressure. However, it is also possible to carry them out under positive pressure or under negative pressure (e.g. in a range from 0.5 to 5 bar).
The reactions can, for example, take place in a temperature range of from 0xc2x0 C. to room temperature and under standard pressure.
The compounds of the general formula (II) are novel and can be prepared (e.g. by analogy with Gazz. Chim. Ital., 97, 1967, 25-33) by converting compounds of the general formula (VII)
R1xe2x80x94CHOxe2x80x83xe2x80x83(VII),
in which
R1 has the abovementioned meaning,
by reaction in the system 1.) CH3xe2x80x94NO2/KF and 2.) K2Cr2O4, into the compounds of the general formula (VIII)
R1xe2x80x94COxe2x80x94CH2xe2x80x94NO2xe2x80x83xe2x80x83(VIII),
in which
R1 has the abovementioned meaning,
subsequently preparing the compounds of the general formula (IX) 
in which
R1 has the abovementioned meaning,
by reaction with hydroxylamine,
cyclizing these compounds with ethyl oxalyl chloride, in a solvent (e.g. ether), to give the compounds of the general formula (X) 
in which
R1 has the abovementioned meaning,
converting these compounds, by the action of ammonia, into the compounds of the general formula (XI) 
in which
R1 has the abovementioned meaning,
and, in a last step, reducing these compounds with Zn/NH4Cl to give the compounds of the general formula (II).
The compounds of the general formula (III) are known.
The compounds of the general formula (IV) are not known.
The compounds of the general formula (V) are not known.
The compounds of the general formula (VI) are not known.
The compounds of the general formula (VII) are known.
The compounds of the general formula (VIII) are known.
The compounds of the general formula (IX) are known.
The compounds of the general formula (X) are known.
The compounds of the general formula (XI) are known.
The compounds according to the invention of the general formula (I) exhibit a valuable pharmacological spectrum of activity which it was not possible to foresee.
They inhibit either one or several of the c-GMP-metabolizing phosphodiesterases (PDE I, PDE II and PDE V). This leads to an increase in c-GMP. The differing expression of the phosphodiesterases in different cells, tissues and organs, as well as the differing subcellular location of these enzymes, make it possible, in combination with the selective inhibitors according to the invention, to address the different cGMP-regulated processes selectively.
In addition, the compounds according to the invention augment the effect of substances such as, for example, EDRF (endothelium-derived relaxing factor) and ANP (atrial natriuretic peptide), of nitro vasodilators and all other substances which increase the concentration of cGMP in another way than phosphodiesterase inhibitors.
The compounds according to the invention of the general formula (I) are therefore suitable for the prophylaxis and/or treatment of diseases in which an increase in the concentration of cGMP is therapeutic, i.e. diseases which are connected with cGMP-regulated processes (usually simply termed xe2x80x9ccGMP-related diseasesxe2x80x9d). These diseases include cardiovascular diseases, diseases of the urogenital system and cerebrovascular diseases.
Within the meaning of the present invention, the term xe2x80x9ccardiovascular diseasesxe2x80x9d covers diseases such as, for example, high blood pressure, neuronal hypertension, stable and unstable angina, peripheral and cardiac vascular diseases, arrhythmias, thromboembolic diseases and ischemias such as myocardial infarction, stroke, transistory and ischemic attacks, angina pectoris, peripheral circulatory disturbances, prevention of restenoses following thrombolysis therapy, percutaneous transluminal angioplasty (PTA), percutaneous transluminal coronary angioplasties (PTCA) and bypass.
Furthermore, the compounds according to the invention of the general formula (I) can also have importance for cerebrovascular diseases. These include, for example, cerebral ischemia, stroke, reperfusion damage, brain trauma, edemas, cerebral thrombosis, dementia and Alzheimer""s disease.
The relaxing effect on smooth musculature makes them suitable for treating disorders of the urogenital system such as prostate hypertrophy and incontinence and also, in particular, for treating erectile dysfunction and female sexual dysfunction.
Activity of the Phosphordiesterases (PDEs)
The cGMP-stimulatable PDE II, the cGMP-inhibitable PDE III and the cAMP-specific PDE IV were isolated either from porcine heart myocardium or from bovine heart myocardium. The Ca+-calmodulin-stimulatable PDE I was isolated from porcine aorta, porcine brain or, preferably, from bovine aorta. The c-GMP-specific PDE V was obtained from porcine small intestine, porcine aorta, human blood platelets and, preferably, from bovine aorta. Purification was effected by means of anion exchange chromatography on Pharmacia MonoQR, essentially in accordance with the method described by M. Hoey and Miles D. Houslay, Biochemical Pharmacology, Vol. 40, 193-202 (1990) and C. Lugman et al., Biochemical Pharmacology, Vol. 35, 1743-1751 (1986).
The enzyme activity is determined in a 100 xcexcl test formulation, in 20 mM Tris/HCl buffer pH 7.5, which contains 5 mM MgCl2, 0.1 mg/ml bovine serum albumin and either 800 Bq of 3HcAMP or 3HcGMP. The final concentration of the corresponding nucleotides is 10xe2x88x926 mol/l. The reaction is started by adding the enzyme, with the quantity of enzyme being measured such that approx. 50% of the substrate is transformed during the incubation time of 30 min. In order to test the cGMP-stimulatable PDE II, 3HcAMP is used as substrate and 10xe2x88x926 mol/l unlabeled cGMP is added to the formulation. In order to test the Ca2+-calmodulin-dependent PDE I, 1 xcexcM CaCl2 and 0,1 xcexcM calmodulin are additionally added to the reaction formulation. The reaction is stopped by adding 100 xcexcl of acetonitrile which contains 1 mM cAMP and 1 mM AMP. 100 xcexcl of the reaction formulation are separated by HPLC and the cleavage products are determined quantitatively online using a flow-through scintillation counter. The substance concentration at which the reaction rate is decreased by 50% is measured. The xe2x80x9cphosphodiesterase [3H] cAMP-SPA enzyme assayxe2x80x9d and the xe2x80x9cphosphodiesterase [3H] cGMP-SPA enzyme assayxe2x80x9d, supplied by Amersham Life Science, were additionally used for the testing. The test was carried out using the experimental protocol specified by the manufacturer. The [3H] cAMP-SPA assay was used for determining the activity of PDE II, with 10xe2x88x926 M cGMP being added to the reaction formulation for the purpose of activating the enzyme. 10xe2x88x927 M calmodulin and 1 xcexcM CaCl2 were added to the reaction formulation for the purpose of measuring PDE I. PDE V was measured using the [3H] cGMP-SPA assay.
In principle, the inhibition of one or more phosphodiesterases of this type leads to an increase in the concentration of cGMP. As a result, the compounds are of interest for all therapies in which an increase in the concentration of cGMP can be assumed to be therapeutic.
The investigation of the cardiovascular effects was carried out on normotensive rats and on SH rats and on dogs. The substances were administered intravenously or orally.
The examination for erection-inducing effect was carried out on conscious rabbits [H. Naganuma, T. Egashira, J. Fuji, Clinical and Experimental Pharmacology and Physiology, 20, 177-183 (1993)]. The substances were administered orally or parenterally.
The novel active compounds, and also their physiologically harmless salts (e.g. hydrochlorides, maleates or lactates), can be converted, in a known manner, into the customary formulations, such as tablets, coated tablets, pills, granules, aerosols, syrups, emulsions, suspensions and solutions, using inert, nontoxic, pharmaceutically suitable carrier substances or solvents. In this connection, the therapeutically effective compound should in each case be present at a concentration of from about 0.5 to 90% by weight of the total mixture, i.e. in quantities which are sufficient for achieving the specified dosage range.
The formulations are prepared, for example, by extending the active compounds with solvents and/or carrier substances, where appropriate using emulsifiers and/or dispersants, with it being possible, for example when using water as diluent, to use organic solvents as auxiliary solvents, where appropriate.
The administration is effected in a customary manner, preferably orally, transdermally or parenterally, for example perlingually, by the buccal route, intravenously, nasally, rectally or by inhalation.
For use in humans, doses of from 0.001 to 50 mg/kg, preferably 0.01 mg/kg-20 mg/kg, are generally administered when administering orally. A dose of 0.001 mg/kg-0.5 mg/kg is expedient when administering parenterally, such as, for example, by way of mucosae, nasally, by the buccal route or by inhalation.
Despite this, it may be necessary, where appropriate, to depart from the mentioned quantities, specifically in dependence on the body weight or the nature of the route of administration, on the individual response to the medicament, on the nature of its formulation and on the time or interval at which the administration takes place. Thus, it can in some cases be sufficient to make do with less than the abovementioned smallest quantity whereas, in other cases, the mentioned upper limit has to be exceeded. When relatively large quantities are being administered, it may be advisable to divide up these quantities into several individual doses which are given during the course of the day.
The compounds according to the invention are also suitable for use in veterinary medicine. For uses in veterinary medicine, the compounds, or their nontoxic salts, can be administered in a suitable formulation, in accordance with common veterinary procedures. The veterinarian can establish the nature of the use, and the dose, in accordance with the nature of the animal to be treated.
Preparing the precursors